Test ID: WARFP
Warfarin Sensitivity Genotype by Sequence Analysis, Blood
Secondary ID 
A test code used for billing and in test definitions created prior to November 2011
NY State Approved Indicates the status of NY State approval and if the test is orderable for NY State clients.
Useful For Suggests clinical disorders or settings where the test may be helpful
Identifying patients who may require warfarin dosing adjustments(2,3) including:
-Patients who have previously been prescribed warfarin and have required multiple dosing adjustments to maintain the international normalized ratio in the target range
-Patients with a history of thrombosis or bleeding when previously taking warfarin
-Patients being started on a first prescription for warfarin
Profile Information A profile is a group of laboratory tests that are ordered and performed together under a single Mayo Test ID. Profile information lists the test performed, inclusive of the test fee, when a profile is ordered and includes reporting names and individual availability.
| Test ID | Reporting Name | Available Separately | Always Performed |
|---|---|---|---|
| WARFS | Warfarin Sensitivity, Genotype | No | Yes |
| WARFQ | Warfarin Sensitivity Genotyping | No | Yes |
Testing Algorithm Delineates situation(s) when tests are added to the initial order. This includes reflex and additional tests.
Special Instructions and Forms Describes specimen collection and preparation information, test algorithms, and other information pertinent to test. Also includes pertinent information and consent forms to be used when requesting a particular test
Method Name A short description of the method used to perform the test
Polymerase Chain Reaction (PCR) Followed by DNA Sequence Analysis
Reporting Name A shorter/abbreviated version of the Published Name for a test; an abbreviated test name
Aliases Lists additional common names for a test, as an aid in searching
Coumadin Genotype
CYP2C9
Cytochrome P450 2C9 Genotyping
P450 2C9 Genotyping
Vitamin K Epoxide Reductase Subunit 1
Vitamin K Genotype (VKORC1)
VKORC1 and 2C9
Warfarin Genotype
Specimen Type Describes the specimen type needed for testing
Specimen Required Defines the optimal specimen. This field describes the type of specimen required to perform the test and the preferred volume to complete testing. The volume allows automated processing, fastest throughput and, when indicated, repeat or reflex testing.
This test is used for assessing CYP2C9 and VKORC1 genes for polymorphisms affecting the metabolism of warfarin/Coumadin. This assay should be ordered on patients who are receiving warfarin for the first time or who require decreased warfarin dosing to maintain the international normalized ratio (INR) in the therapeutic range.
If using drugs other than warfarin, order 2C9S/60528 Cytochrome P450 2C9 Genotype by Sequence Analysis, Blood which only includes testing for the CYP2C9 gene.
Multiple whole blood EDTA genotype tests can be performed on a single specimen after a single extraction. See Multiple Whole Blood EDTA Genotype Tests in Special Instructions for a list of tests that can be ordered together.
Container/Tube: Lavender top (EDTA)
Specimen Volume: 3 mL
Collection Instructions: Send specimen in original tube.
Additional Information:
1. Bone marrow and liver transplants will interfere with testing. Call Mayo Medical Laboratories at 800-533-1710 or 507-266-5700 for instructions.
2. Transfusions will interfere with testing for up to 4 to 6 weeks. DNA obtained from white cells may not provide useful information for patients who received a recent transfusion of blood that was not leukocyte-reduced. Wait 4 to 6 weeks until transfused cells have left the patient's circulation before drawing the patient's blood specimen for genotype testing.
3. Cytochrome P450 Patient Education Brochure (Supply T526) is available upon request.
Forms: New York Clients-Informed consent is required. Please document on the request form or electronic order that a copy is on file. An Informed Consent for Genetic Testing (Supply T576) is available in Special Instructions.
Specimen Minimum Volume Defines the amount of specimen required to perform an assay once, including instrument and container dead space. Submitting the minimum specimen volume makes it impossible to repeat the test or perform confirmatory or perform reflex testing. In some situations, a minimum specimen volume may result in a QNS (quantity not sufficient) result, requiring a second specimen to be collected.
Reject Due To Identifies specimen types and conditions that may cause the specimen to be rejected
| Hemolysis | NA |
| Lipemia | NA |
| Icterus | NA |
| Other | NA |
Specimen Stability Information Provides a description of the temperatures required to transport a specimen to the laboratory. Alternate acceptable temperature(s) are also included.
| Specimen Type | Temperature | Time |
|---|---|---|
| Whole Blood EDTA | Ambient (preferred) | |
| Refrigerated | ||
Clinical Information Discusses physiology, pathophysiology, and general clinical aspects, as they relate to a laboratory test
Warfarin is a coumarin-based drug commonly utilized in anticoagulation therapy to prevent thrombosis due to inherited and acquired hemostatic disorders. The drug is also used in a number of other medical conditions and treatments including atrial fibrillation and hip replacement surgery. Warfarin acts by interfacing with the metabolism of vitamin K, which is necessary for production of key coagulation factors. Warfarin inhibits vitamin K recycling by blocking its metabolism at the vitamin K-epoxide intermediate thereby decreasing the amount of available vitamin K. Warfarin has a narrow therapeutic window; under medicating increases the risk for thrombosis and overmedicating increases the risk for cerebrovascular accidents. Warfarin therapy has one of the highest rates of severe adverse drug reactions.
Warfarin is dosed using non genetic factors including gender, weight, and age, and is monitored by coagulation testing in order to maintain the international normalized ratio (INR) within specific limits. However, warfarin metabolism is highly variable and dependent upon genetic factors. Polymorphisms within 2 genes are known to affect the metabolism of warfarin and the dose needed to maintain the correct serum drug level and degree of anticoagulation, as measured by the INR.
The cytochrome P450 2C9 gene (CYP2C9) encodes an enzyme that metabolizes the more active isomer of warfarin (S-warfarin) to inactive products. Polymorphisms in this gene decrease the activity of the enzyme and may cause increases in serum warfarin and overmedicating, driving INR above the therapeutic target level.
The second gene (VKORC1) encodes vitamin K epoxide reductase complex subunit-1 (VKORC1), a small transmembrane protein of the endoplasmic reticulum that is part of the vitamin K cycle and the target of warfarin therapy.(1) VKORC1 is primarily transcribed in the liver, although it is present in smaller amounts in the heart and pancreas. Vitamin K epoxide, a by-product of the carboxylation of blood coagulation factors, is reduced to vitamin K by VKORC1. A polymorphism within the promoter of VKORC1 decreases expression of the gene, decreasing the availability of vitamin K. This may cause increases in serum warfarin and overmedicating, driving INR above the therapeutic target level.
Thus, in both situations (polymorphisms in either CYP2C9 or VKORC1), a reduced warfarin dose is needed to compensate for the effects of the polymorphism in order to maintain the target INR.
CYP2C9
CYP2C9 metabolizes a wide variety of drugs including warfarin and many nonsteroidal anti-inflammatory drugs. It is also partially responsible for metabolizing other drugs such as fluoxetine, fluvastatin, phenytoin, and oral hypoglycemic drugs.
A number of specific polymorphisms have been found in the CYP2C9 gene that result in enzymatic deficiencies. The following information outlines the relationship between the polymorphisms detected in this assay and the effect on the activity of the enzyme encoded by that allele:
| CYP2C9 Allele | Nucleotide Change | Effect on Enzyme Metabolism |
| *1 | None (wild type) | Extensive metabolizer (normal) |
| *2 | 430C->T | Reduced activity |
| *3 | 1075A->C | Minimal activity |
| *4 | 1076T->C | Reduced activity |
| *5 | 1080C->G | Reduced activity |
| *6 | 818delA | No activity |
Dosing of warfarin, which is metabolized through CYP2C9, may require adjustment for the individual patient. Patients who are poor metabolizers (reduced activity) may benefit by dose reductions or by being switched to other comparable drugs that are not metabolized primarily by CYP2C9. The following is a partial listing of drugs known to affect CYP2C9 activity as of the date of this report. A more complete listing is presented in the drug label, available at URL:
http://www.accessdata.fda.gov/drugsatfda_docs/label/2010/009218s108lbl.pdf
Drugs that undergo metabolism by CYP2C9:
-Angiotensin II blockers: irbesartan, losartan
-Anticoagulants: warfarin (more active S-isomer)
-Antidepressants: amitriptyline (minor), fluoxetine
-Nonsteroidal anti-inflammatory drugs (NSAIDS): celecoxib, diclofenac, ibuprofen, naproxen, piroxicam, suprofen
-Oral hypoglycemic agents: glipizide, glimepiride, glyburide/glibenclamide, nateglinide, tolbutamide
-Miscellaneous drugs: fluvastatin, phenytoin, sulfamethoxazole, tamoxifen, torsemide
-Coadministration of these drugs may decrease the rate of elimination of other drugs metabolized by CYP2C9
Drugs known to increase CYP2C9 activity:
-Phenobarbital, rifampin, secobarbital
-Coadministration of these drugs increase the synthesis of CYP2C9, resulting in increased CYP2C9 activity and metabolism of warfarin. A dose increase may be needed to maintain the INR in the target range
Drugs known to decrease CYP2C9 activity:
-Amiodarone, fenofibrate, fluconazole, fluvastatin, isoniazid, lovastatin, phenylbutazone, sertraline, sulfamethoxazole, sulfaphenazole, teniposide, ticlopidine, voriconazole, zafirlukast
-Coadministration of these drugs may decrease the rate of metabolism of CYP2C9-metabolized drugs, including warfarin, increasing the possibility of toxicity
VKORC1
The -1639 promoter polymorphism is located in the second nucleotide of an E-Box (CANNTG) and its presence disrupts the consensus sequence, reducing promoter activity. In vitro experiments show a 44% higher transcription level of the G versus the A allele.(1) The -1639 G->A nucleotide change results in decreased gene expression and reduced enzyme activity.
Reference Values Describes reference intervals and additional information for interpretation of test results. May include intervals based on age and sex when appropriate. Intervals are Mayo-derived, unless otherwise designated. If an interpretive report is provided, the reference value field will state this.
An interpretive report will be provided.
Interpretation Provides information to assist in interpretation of the test results
An interpretive report will be provided.
The normal genotype (wild-type) for CYP2C9 is termed CYP2C9*1. Other genotypes that lead to inactive or reduced activity alleles include CYP2C9*2, CYP2C9*3, CYP2C9*4, CYP2C9*5, and CYP2C9*6. An individual who has homozygous wild-type, CYP2C9*1/CYP2C9*1, is considered an extensive metabolizer.
The normal genotype for VKORC1 is -1639G. A polymorphism at -1639A reduces VKORC1 expression. The VKORC1 GA or AA genotype leads to a significant decrease in mRNA expression in the liver compared with individuals with the GG genotype.
Individuals who have polymorphisms in both the VKORC1 promoter (GA or AA) and also in CYP2C9 should receive a reduced dose of warfarin to maintain the international normalized ratio in the target range; dosing adjustments are required when polymorphisms in both genes are present.
Drug-drug interactions and drug-metabolite inhibition must be considered when dealing with heterozygous individuals. Drug-metabolite inhibition can occur, resulting in inhibition of residual functional CYP2C9 or VKORC1 catalytic activity. A clinical pharmacologist should be consulted for assessing the potential for drug interactions.
Patients may also develop toxicity problems if liver and kidney function are impaired.
Cautions Discusses conditions that may cause diagnostic confusion, including improper specimen collection and handling, inappropriate test selection, and interfering substances
This assay should be ordered on patients who require decreased warfarin dosing to maintain the international normalized ratio in the therapeutic range.
This test detects only the specified polymorphisms. Additional findings, such as small insertions and deletions or novel mutations, will be reported if found. Other polymorphisms in the primer binding regions can affect the testing, and, ultimately, the genotyping assessments made.
Warfarin metabolism may be inhibited through drug-drug interactions, including amiodarone and statins.
Genotyping patients using DNA obtained from leukocytes may not provide useful information in patients who have had a bone marrow or liver transplant or a recent transfusion. To obtain an accurate genotype on a bone marrow transplant recipient, buccal cells should be provided. To obtain an accurate genotype for a patient who has received a donor liver, testing must be done on donor cells. If the patient has been transfused, wait 4 to 6 weeks until transfused cells have left the circulation.
Clinical Reference Provides recommendations for further in-depth reading of a clinical nature
1. Oldenburg J, Bevens C, Muller C, Watzka M: Vitamin K epoxide reductase complex subunit I (VKORC1): the key protein of the vitamin K cycle. Antioxid Redox Signal 2006;8(3-4):347-353
2. Yuan H, Chen J, Lee M, et al: A novel functional VKORC1 promoter polymorphism is associated with inter-individual and inter-ethnic differences in warfarin sensitivity. Hum Mol Genet 2005;14:1745-1751
3. Sconce E, Khan T, Wynne H, et al: The impact of CYP2C9 and VKORC1 genetic polymorphism and patient characteristics upon warfarin dose requirements proposal for a new dosing regimen. Blood 2005;106:2329-2333
Method Description Describes how the test is performed and provides a method-specific reference
Genomic DNA is extracted from whole blood. The CYP2C9 gene and VKORC1 promoter are amplified by PCR. The PCR product is then purified and sequenced in both directions using fluorescent dye-terminator chemistry. Sequencing products are separated on an automated sequencer and trace files analyzed for variations in exons 3, 5, and 7 of the CYP2C9 gene and the VKORC1 gene promoter using mutation detection software and visual inspection. (Unpublished Mayo method)
Day(s) and Time(s) Test Performed Outlines the days and times the test is performed. This field reflects the day and time the sample must be in the testing laboratory to begin the testing process and includes any specimen preparation and processing time required before the test is performed. Some tests are listed as continuously performed, which means assays are performed several times during the day.
Varies
Analytic Time Defines the amount of time it takes the laboratory to setup and perform the test. This is defined in number of days. The shortest interval of time expressed is "same day/1 day," which means the results may be available the same day that the sample is received in the testing laboratory. One day means results are available 1 day after the sample is received in the laboratory.
Maximum Laboratory Time Defines the maximum time from specimen receipt at Mayo Medical Laboratories until the release of the test result
Specimen Retention Time Outlines the length of time after testing that a specimen is kept in the laboratory before it is discarded
Performing Laboratory Location The location of the laboratory that performs the test
Test Classification Provides information regarding the medical device classification for laboratory test kits and reagents. Tests may be classified as cleared or approved by the US Food and Drug Administration (FDA) and used per manufacturer's instructions, or as products that do not undergo full FDA review and approval, and are then labeled as an Analyte Specific Reagent (ASR), Investigation Use Only (IUO) product, or a Research Use Only (RUO) product.
CPT Code Information Provides guidance in determining the appropriate Current Procedural Terminology (CPT) code(s) information for each test or profile. The listed CPT codes reflect Mayo Medical Laboratories interpretation of CPT coding requirements. It is the responsibility of each laboratory to determine correct CPT codes to use for billing.
81227-CYP2C9 (cytochrome P450, family 2, subfamily C, polypeptide 9) (eg, drug metabolism), gene analysis, common variants (eg, *2, *3, *5, *6)
81355-VKORC1 (vitamin K epoxide reductase complex, subunit 1) (eg, warfarin metabolism), gene analysis, common variants (eg, -1639/3673)
LOINC® Code Information Provides guidance in determining the Logical Observation Identifiers Names and Codes (LOINC) values for the result codes returned for this test or profile.
| Result ID | Reporting Name | LOINC Code |
|---|---|---|
| WARFQ | Warfarin Sensitivity Genotyping | In Process |
| 26941 | Warfarin Sens Phenotype Interp | 69047-9 |
| 32183 | CYP2C9 Star Alleles/VKORC1 Genotype | 54451-0 |
| 28359 | CYP2C9 430C>T(*2) | In Process |
| 28360 | CYP2C9 818delA(*6) | In Process |
| 28361 | CYP2C9 1075A>C(*3) | In Process |
| 28362 | CYP2C9 1076T>C(*4) | In Process |
| 28363 | CYP2C9 1080C>G(*5) | In Process |
| 87988 | VKORC1 -1639G>A | In Process |
| 28332 | Warfarin Sens Genotype Interp | 69047-9 |
| 28364 | Warfarin Sensitivity Reviewed by | In Process |
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